Presently, all attempts at cellular-based tissue engineering are being done at the manufacturing site where uniform and continuous infiltration of cells into a matrix are prepared together before shipping for implantation. The manufacturing of cellular based medical devices is costly and time consuming. Part of the lengthy time consumption results from the necessity of incubating the cells in the matrix for long periods before in vivo use. For example, in the overview of a company involved in the preparation of cell scaffolds, it is stated that “after [the cells] develop over a period of a few weeks, the cells on the scaffold are kept under optimal conditions in bioreactors.
The bioreactors are challenging to build because they have to create perfect conditions to grow the implants.” (Advanced Tissue Sciences, Inc., La Jolla, Calif.).
It is known that cellular tissue engineering requires continuous biological devices that are pre-constructed prior to implantation (L. Germain et al., 2000, Medical & Biological Engineering & Computing, 38(2):232-40; W. W. Minuth et al., 1998, Cell Tissue Res., 291(1):1-11; G. K. Naughton et al., 1999, Clin. in Plastic Surgery, 26(4):579-86, viii). Until the present time using the invention described herein, most cell-matrices were made using non-stem cells. However, these cells have proven to be difficult to work with (F. Berthod and O. Damour, 1997, British Journal of Dermatology, 136:809-816). In order to produce such non-stem cell matrices, prolonged incubation times are needed after the cells have been incorporated into the matrix. As a particular example, it has been reported that at least 3 to 5 hours of incubation time is needed for fibroblasts to attach to a scaffold and another 2 to 3 weeks of incubation time is required to achieve confluence (J. F. Hansbrough et al., 1991, Surgery, 111(4):438-446). Moreover, the use of collagen sponges requires that the cells have to be incubated in the sponge for at least 24 hours, and another 8 to 10 days are needed for the cells to achieve confluence (F. Berthod et al., 1993, Biomaterials, 14(10): 749-754).
In addition, currently available cell-matrices require careful monitoring. Cells on scaffolds are vigorously tested to confirm and maintain the proper metabolic rate that will allow the cells on scaffolds to grow and reproduce in a steady-state manner resulting in optimal materials (Advanced Tissue Sciences, Inc. La Jolla, Calif.).
For cell matrix products used in wound coverage, there has been a perpetual belief that uniform layers of continuous cells are needed for a functional product (F. A. Auger et al., 1988, Med. Biol. Eng. Compu., 36: 801-812; S. T. Boyce, 1996, Tissue Eng., 2: 255-266; O. Damour, et al., 1997, “Cultured autologous epidermis for massive burn wounds: 15 years of practice”. Rouabhia, M. (Ed.): Skin substitute production by tissue engineering: clinical and fundamental applications. Landes, Austin, pp. 23-45). As an example, the importance of continuous layers has been emphasized in conjunction with the marketing of the product Apligraf® (Organogenesis, Inc., Canton, Mass.), which is carefully cultured on scaffold in a bilayer for over 20 days to produce a continuous cell layer of dermis and epidermis.
Current methods of producing cell matrices for in vivo tissue and organ repair are very costly and time consuming. Such cell matrices are costly due to the specialized factories and/or procedures needed to produce these products. Also, since cell-matrix products involve living biological cells/tissue, a tremendous loss of product occurs from shipping, the delays associated therewith, and the like. Additionally, given the nature of the products, obtaining regulatory approval for new products that are based on living cells and a new matrix poses difficulties.
Thus, there is a serious need for cell-matrix compositions that are low in cost, that are versatile, and easily prepared and/or manufactured. There is a further need for cell matrix compositions that do not require extensive in vitro incubation or cultivation periods after the cells have been incorporated into the matrix. Those in the art have recognized that a major problem remaining to be solved is the delay in producing the cell-matrix product after initial preparation. Specifically, it has been stated that there is a problem of a three week delay necessary to produce a sufficient amount of autologous keratinocytes and fibroblasts for the production of reconstructed skin. (F. Berthod and O. Damour, 1997, British Journal of Dermatology, 136: 809-816). The present invention provides a solution for the above-mentioned problems and delays currently extant in the art.